RESUMEN
Cardiac troponin I (cTnI) is a key regulator of cardiomyocyte contraction. However, its role in mitochondria is unknown. Here we show that cTnI localized to mitochondria in the heart, inhibited mitochondrial functions when stably expressed in noncardiac cells and increased the opening of the mitochondrial permeability transition pore under oxidative stress. Direct, specific and saturable binding of cTnI to F1FO-ATP synthase was demonstrated in vitro using immune-captured ATP synthase and in cells using proximity ligation assay. cTnI binding doubled ATPase activity, whereas skeletal troponin I and several human pathogenic cTnI variants associated with familial hypertrophic cardiomyopathy did not. A rationally designed peptide, P888, inhibited cTnI binding to ATP synthase, inhibited cTnI-induced increase in ATPase activity in vitro and reduced cardiac injury following transient ischemia in vivo. We suggest that cTnI-bound ATP synthase results in lower ATP levels, and releasing this interaction during cardiac ischemia-reperfusion may increase the reservoir of functional mitochondria to reduce cardiac injury.
Asunto(s)
Mitocondrias Cardíacas , ATPasas de Translocación de Protón Mitocondriales , Troponina I , Animales , Humanos , Masculino , Ratones , Ratas , Adenosina Trifosfato/metabolismo , Modelos Animales de Enfermedad , Células HEK293 , Ratones Endogámicos C57BL , Mitocondrias Cardíacas/metabolismo , Poro de Transición de la Permeabilidad Mitocondrial/metabolismo , ATPasas de Translocación de Protón Mitocondriales/metabolismo , Isquemia Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Estrés Oxidativo/efectos de los fármacos , Unión Proteica , Troponina I/metabolismoRESUMEN
COVID-19 emergency use authorizations and approvals for vaccines were achieved in record time. However, there remains a need to develop additional safe, effective, easy-to-produce, and inexpensive prevention to reduce the risk of acquiring SARS-CoV-2 infection. This need is due to difficulties in vaccine manufacturing and distribution, vaccine hesitancy, and, critically, the increased prevalence of SARS-CoV-2 variants with greater contagiousness or reduced sensitivity to immunity. Antibodies from eggs of hens (immunoglobulin Y; IgY) that were administered the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein were developed for use as nasal drops to capture the virus on the nasal mucosa. Although initially raised against the 2019 novel coronavirus index strain (2019-nCoV), these anti-SARS-CoV-2 RBD IgY surprisingly had indistinguishable enzyme-linked immunosorbent assay binding against variants of concern that have emerged, including Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617.2), and Omicron (B.1.1.529). This is different from sera of immunized or convalescent patients. Culture neutralization titers against available Alpha, Beta, and Delta were also indistinguishable from the index SARS-CoV-2 strain. Efforts to develop these IgY for clinical use demonstrated that the intranasal anti-SARS-CoV-2 RBD IgY preparation showed no binding (cross-reactivity) to a variety of human tissues and had an excellent safety profile in rats following 28-day intranasal delivery of the formulated IgY. A double-blind, randomized, placebo-controlled phase 1 study evaluating single-ascending and multiple doses of anti-SARS-CoV-2 RBD IgY administered intranasally for 14 days in 48 healthy adults also demonstrated an excellent safety and tolerability profile, and no evidence of systemic absorption. As these antiviral IgY have broad selectivity against many variants of concern, are fast to produce, and are a low-cost product, their use as prophylaxis to reduce SARS-CoV-2 viral transmission warrants further evaluation. Clinical Trial Registration: https://www.clinicaltrials.gov/ct2/show/NCT04567810, identifier NCT04567810.
Asunto(s)
COVID-19 , SARS-CoV-2 , Animales , Anticuerpos Antivirales , COVID-19/prevención & control , Pollos , Femenino , Humanos , Inmunoglobulinas , Ratas , Glicoproteína de la Espiga del CoronavirusRESUMEN
The ALDH2*2 missense variant that commonly causes alcohol flushing reactions is the single genetic polymorphism associated with the largest number of traits in humans. The dysfunctional ALDH2 variant affects nearly 8% of the world population and is highly concentrated among East Asians. Carriers of the ALDH2*2 variant commonly present alterations in a number of blood biomarkers, clinical measurements, biometrics, drug prescriptions, dietary habits and lifestyle behaviors, and they are also more susceptible to aldehyde-associated diseases, such as cancer and cardiovascular disease. However, the interaction between alcohol and ALDH2-related pathology is not clearly delineated. Furthermore, genetic evidence indicates that the ALDH2*2 variant has been favorably selected for in the past 2000-3000â years. It is therefore necessary to consider the disease risk and mechanism associated with ALDH2 deficiency, and to understand the possible beneficial or protective effect conferred by ALDH2 deficiency and whether the pleiotropic effects of ALDH2 variance are all mediated by alcohol use.
Asunto(s)
Consumo de Bebidas Alcohólicas , Polimorfismo Genético , Consumo de Bebidas Alcohólicas/genética , Aldehído Deshidrogenasa Mitocondrial/genética , Pueblo Asiatico , Etanol , HumanosRESUMEN
Human aldehyde dehydrogenase (ALDH) is a multigene family with 19 functional members encoding a class of diverse but important enzymes for detoxification or biotransformation of different endogenous and exogenous aldehyde substrates. Genetic mutations in the ALDH genes can cause the accumulation of toxic aldehydes and abnormal carbonyl metabolism and serious human pathologies. However, the physiological functions and substrate specificity of many ALDH genes are still unknown. Although many genetic variants of the ALDH gene family exist in human populations, their phenotype or clinical consequences have not been determined. Using the most comprehensive global human Genome Aggregation Database, gnomAD, we annotated here 1350 common variants in the 19 ALDH genes. These 1350 common variants represent all known genetic polymorphisms with a variant allele frequency of ≥0.1% (or an expected occurrence of ≥1 carrier per 500 individuals) in any of the seven major ethnic groups recorded by gnomAD. We detailed 13 types of DNA sequence variants, their genomic positions, SNP ID numbers, and allele frequencies among the seven major ethnic groups worldwide for each of the 19 ALDH genes. For the 313 missense variants identified in the gnomAD, we used two software algorithms, Polymorphism Phenotyping (PolyPhen) and Sorting Intolerant From Tolerant (SIFT), to predict the consequences of the variants on the structure and function of the enzyme. Finally, gene constraint analysis was used to predict how well genetic mutations were tolerated by selection forces for each of the ALDH genes in humans. Based on the ratio of observed and expected variant numbers in gnomAD, the three ALDH1A gene members, ALDH1A1, ALDH1A2, and ALDH1A3, appeared to have the lowest tolerance for loss-of-function mutations as compared to the other ALDH genes (# observed/# expected ratio 0.15-0.26). These analyses suggest that the ALDH1A1, ALDH1A2, and ALDH1A3 enzymes may serve a more essential function as compared with the other ALDH enzymes; functional loss mutations are much less common in healthy human populations than expected. This informatic analysis may assist the research community in determining the physiological function of ALDH isozymes and associate common variants with clinical phenotypes.
Asunto(s)
Aldehído Deshidrogenasa/genética , Genoma Humano/genética , Genómica , Familia de Multigenes/genética , Aldehído Deshidrogenasa/clasificación , Bases de Datos Genéticas , Frecuencia de los Genes/genética , Humanos , Anotación de Secuencia Molecular , Polimorfismo Genético/genéticaRESUMEN
In December 2019, a novel coronavirus, termed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was identified as the cause of pneumonia with severe respiratory distress and outbreaks in Wuhan, China. The rapid and global spread of SARS-CoV-2 resulted in the coronavirus 2019 (COVID-19) pandemic. Earlier during the pandemic, there were limited genetic viral variations. As millions of people became infected, multiple single amino acid substitutions emerged. Many of these substitutions have no consequences. However, some of the new variants show a greater infection rate, more severe disease, and reduced sensitivity to current prophylaxes and treatments. Of particular importance in SARS-CoV-2 transmission are mutations that occur in the Spike (S) protein, the protein on the viral outer envelope that binds to the human angiotensin-converting enzyme receptor (hACE2). Here, we conducted a comprehensive analysis of 441,168 individual virus sequences isolated from humans throughout the world. From the individual sequences, we identified 3540 unique amino acid substitutions in the S protein. Analysis of these different variants in the S protein pinpointed important functional and structural sites in the protein. This information may guide the development of effective vaccines and therapeutics to help arrest the spread of the COVID-19 pandemic.
Asunto(s)
Variación Genética , Glicoproteína de la Espiga del Coronavirus/genética , Sustitución de Aminoácidos , Furina/metabolismo , Glicosilación , Humanos , Modelos Moleculares , Profilaxis Pre-Exposición , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/aislamiento & purificaciónRESUMEN
A mutation analysis of SARS-CoV-2 genomes collected around the world sorted by sequence, date, geographic location, and species has revealed a large number of variants from the initial reference sequence in Wuhan. This analysis also reveals that humans infected with SARS-CoV-2 have infected mink populations in the Netherlands, Denmark, United States, and Canada. In these animals, a small set of mutations in the spike protein receptor binding domain (RBD), often occurring in specific combinations, has transferred back into humans. The viral genomic mutations in minks observed in the Netherlands and Denmark show the potential for new mutations on the SARS-CoV-2 spike protein RBD to be introduced into humans by zoonotic transfer. Our data suggests that close attention to viral transfer from humans to farm animals and pets will be required to prevent build-up of a viral reservoir for potential future zoonotic transfer.
Asunto(s)
Visón/virología , Mutación , SARS-CoV-2/genética , Glicoproteína de la Espiga del Coronavirus/genética , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Sitios de Unión , COVID-19/transmisión , COVID-19/veterinaria , COVID-19/virología , Canadá , Dinamarca , Humanos , Países Bajos , Filogenia , SARS-CoV-2/aislamiento & purificación , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/metabolismo , Estados UnidosRESUMEN
Two SARS-CoV-2 variants of concern showing increased transmissibility relative to the Wuhan virus have recently been identified. Although neither variant appears to cause more severe illness nor increased risk of death, the faster spread of the virus is a major threat. Using computational tools, we found that the new SARS-CoV-2 variants may acquire an increased transmissibility by increasing the propensity of its spike protein to expose the receptor binding domain via proteolysis, perhaps by neutrophil elastase and/or via reduced intramolecular interactions that contribute to the stability of the closed conformation of spike protein. This information leads to the identification of potential treatments to avert the imminent threat of these more transmittable SARS-CoV-2 variants.
Asunto(s)
Elastasa Pancreática/metabolismo , SARS-CoV-2/metabolismo , Glicoproteína de la Espiga del Coronavirus/metabolismo , Secuencia de Aminoácidos , Enzima Convertidora de Angiotensina 2/química , Enzima Convertidora de Angiotensina 2/metabolismo , Anticuerpos Neutralizantes/inmunología , COVID-19/patología , COVID-19/virología , Humanos , Simulación de Dinámica Molecular , Mutación , Neutrófilos/citología , Neutrófilos/metabolismo , Unión Proteica , Estabilidad Proteica , Estructura Terciaria de Proteína , SARS-CoV-2/aislamiento & purificación , SARS-CoV-2/patogenicidad , Alineación de Secuencia , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/genética , Glicoproteína de la Espiga del Coronavirus/inmunologíaRESUMEN
Targeted drug delivery to pancreatic islet ß cells is an unmet clinical need. ß cells possess a uniquely high Zn2+ concentration, and integrating Zn2+-binding activity into a small molecule can bias drug accumulation and activity toward ß cells. This protocol can be used to evaluate a molecule's capacity to chelate islet Zn2+, accumulate in islets, and stimulate ß cell-selective replication in mouse pancreas. One obstacle is establishing an LC-MS/MS-based method for compound measurement. Limitations include target compound ionizability and the time-sensitive nature of some experimental assay steps. For complete details on the use and execution of this protocol, please refer to Horton et al. (2019).